Antibacterial silver-doped bioactive silica gel production ... · ORIGINAL RESEARCH Antibacterial...
Transcript of Antibacterial silver-doped bioactive silica gel production ... · ORIGINAL RESEARCH Antibacterial...
ORIGINAL RESEARCH
Antibacterial silver-doped bioactive silica gel productionusing molten salt method
Roya Payami1 • Mohammad Ghorbanpour2 • Aiyoub Parchehbaf Jadid3
Received: 7 February 2016 / Accepted: 11 April 2016 / Published online: 25 April 2016
� The Author(s) 2016. This article is published with open access at Springerlink.com
Abstract Due to broad-spectrum antimicrobial activity,
silver nanoparticles have great application potential in
disinfection of contaminated water. The aim of this
research was the introduction of a fast and simple method
titled as ‘‘molten salt method’’ for the production of silver-
doped bioactive silica gel (SG) nanocomposite. In this
method, SG was imposed into the molten salt of silver
nitrate at 150 and 300 �C for various times. Interestingly,
molten salt method was not utilized any reducing reagent
or other chemicals unless molten silver nitrate. The syn-
thesis and fixing of nanoparticles into the support were
done in \60 min. The prepared silver/SG nanocomposite
was evaluated using scanning electron microscope (SEM),
energy dispersive X-ray fluorescence, leaching test and
antibacterial test. SEM images showed that the contact of
SG with the molten salt caused the formation of nanopar-
ticles on the SG. On the other hand, increasing the contact
time, it led to a larger and increased number of particles.
The antibacterial tests demonstrated that this composite is
suitable for using as antibacterial material. The test of
elution with water indicated that the prepared nanocom-
posite is stable and the amount of the released silver in the
water was negligible.
Keywords Antibacterial � Silver � Silica gel � Molten salt
method
Introduction
Silver nanoparticles have recently attracted great interest
due to their distinctive properties such as large surface
areas, unique physical, chemical and biological properties
[1, 2]. It is widely known that materials containing silver
show antibacterial property [3, 4]. Now, the silver
nanoparticles are emerging as a new generation of
antibacterial agent, which has been used in medical
applications and antibacterial water filter [5]. The use of
silver nanoparticles is particularly potential to treat drink-
ing water, which is frequently infected by antibiotic
resistant bacteria. Treatment of this water usually requires
using high concentrated chlorine compounds, which may
cause a high risk of human cancer [6].
However, aggregation of silver nanoparticles and
leaching of Ag? ions in aquatic system restricted the
application of silver nanoparticles. Therefore, demand for
the production of solid supported silver nanomaterials has
been increased. Generally, the nanoparticle-doped materi-
als have several advantages, such as high performance, low
price (compared to pure silver), high chemical durability
and low release silver ions for a long period of time [7, 8].
There are several methods for preparing silver-doped silica
including multi-target sputtering [2, 9], sol-gel process [4]
and ion exchange process [10, 11]. For example, spherical
nanoparticles with a silver core and an amorphous silica
shell were successfully fabricated using tetraethoxysilane
as silica precursor and reducing silver nitrate with ascorbic
acid. These nanoparticles had excellent antibacterial effects
against E. coli and S. aureus [12]. In a similar approach,
& Mohammad Ghorbanpour
1 Department of Food Science, Sarab Branch, Islamic Azad
University, Sarab, Iran
2 Chemical Engineering Department, University of Mohaghegh
Ardabili, Ardabil, Iran
3 Department of Chemistry, Ardabil Branch, Islamic Azad
University, Ardabil, Iran
123
J Nanostruct Chem (2016) 6:215–221
DOI 10.1007/s40097-016-0193-2
silver nanoparticles were immobilized onto the surface of
magnetic silica composite to prepare magnetic disinfectant
that exhibited enhanced stability and antibacterial activity
[13].
However, the synthesis and fabrication of supported
silver nanomaterial via these methods require extensive
use of toxic and highly cost chemicals and organic
solvents, which frequently raise health and safety issues
[14]. This has led to increasing interest on the devel-
opment of inexpensive preparation methods suitable for
large economic industrial applications. This may sig-
nificantly boost the industrial production of the inex-
pensive silver-doped silica products for various
applications.
In this work, silver-doped bioactive silica gel (SG) was
prepared by a new approach titled as ‘‘molten salt
method’’. To optimize the process condition while guar-
anteeing the antibacterial action, different synthesis con-
ditions (time and temperature) were selected. The obtained
samples were characterized by means of scanning electron
microscope (SEM) and energy dispersive X-ray fluores-
cence (l-EDXRF) observation. The effect of surface
modifications on the antibacterial activity of samples has
been investigated.
Methods
Materials
AgNO3, SG (Kieselgel 60, 0.063–0.200 mm), Mueller–
Hinton broth and nutrient agar were purchased from the
Merck Company (Tehran, Iran). All reagents were used
without further purification. The bacterial strain used for
the antibacterial activity was Gram-negative E. coli (PTCC
1270) and Gram-positive S. aureus (PTCC 1112) received
from the Iranian Research Organization for Science and
Technology.
Preparation of Ag/SG nanocomposite
This process was performed in the molten salt bath of
AgNO3 for different periods ranging from 1 to 30 min.
AgNO3, after weighting, were grounded to get a homoge-
neous mixture. The temperature of the bath was maintained
at about 150 and 300 �C. A quartz beaker was partially
filled with the mixture of silver nitrate (AgNO3) and SG
(1:1, wt/wt), and placed in a furnace which was electrically
heated up to the temperature of the process. The samples
were then taken out of the molten beaker, and were cooled
in air. Later, they were ultrasonically cleaned with distilled
water. After molten salt process, a slight yellow coloration
was observed.
Characterizations
The microstructures of the samples were observed by SEM
(LEO 1430VP, Germany). l-EDXRF analysis was per-
formed using a XMF-104 X-ray Microanalyzer (Unisantis
S.A., Switzerland) equipped with a 50 W molybdenum
tube and a high resolution two-stage Peltier-cooled Si-PIN
detector. The samples were positioned in definite places
and at a constant height of the holder base. The temperature
was controlled between 32 and 34 �C throughout the
experiments. The voltage and current were 30 kV and
300 lA, respectively. Each l-EDXRF analysis was per-
formed in 50 s to obtain sufficient counts. For homogeneity
tests, three different points of approximate constant ori-
entation with respect to the nanocomposites were analyzed
[15].
Water elution test
For each composite, 0.2 g of composite was immersed in
10 mL of distilled water and vigorous agitation in a
shaking water bath (30 �C, 200 RPM) for 2, 6 and 24 h.
Supernatants from each test tube were collected by cen-
trifugation at 4,000 RPM for 10 min. Silver ions released
from the nanocomposites were qualitatively determined by
an atomic absorption spectrometer (AA800, Perkin Elmer).
Water elution test was replicated twice.
Antibacterial activity
The antibacterial activity of the composites against both
E. coli (Gram-negative) and S. aureus (Gram-positive) was
tested by agar diffusion test. Samples were exposed to
bacteria in solid media (nutrient agar), and the inhibition
zone around each sample was measured and recorded as
the antibacterial effect of composites. Agar plates were
inoculated with 100 lL suspensions of bacteria. The
composites and parent SG were placed on agar disks and
incubated at 37 �C for 24 h. The inhibition zone was
measured at three different points. Antibacterial activity
test was replicated twice.
Results
Characterization
After molten salt process, the color of the SG surface
changed from pale yellow to bright yellow, amber and dark
amber depending on the processing time and temperature,
as shown in Fig. 1. For high temperature process, the
coloring becomes darker. The coloration of the sample
prepared at 150 �C for 30 min matches with the color of
216 J Nanostruct Chem (2016) 6:215–221
123
the sample processed at 300 �C for 5 min. The colors of the
products obtained in this work were similar to previous
reports [4, 11].
The un-embedded SG as well as the Ag? appears col-
orless. On the other hand, Ag0 is yellow or brown/gray
depending on the concentration and size of silver
nanoparticles in the silica matrix [4]. The appearance and
disappearance of colors is claimed be associated with the
change in the state of silver (Ag0 or Ag?) at the various
preparation time and temperatures.
Figure 2 shows the morphology of producing SG and
composites. One understands that there is no remarkable
change on the surface after coating with silver when
compared SG surface (Fig. 2b) with produced composites
(Figs. 2c, e). The only difference is the developmental
changes in the micro pores on the initial structure of SG. It
seems that these pores have been filled during the molten
salt process with very small particles of silver. For further
investigation, composites were heated at 100 �C for 3 h.
Based on the available reports, heating causes the move-
ment of silver particles on the support and consequently
their adhesion. The result of this action is the formation of
larger particles [16]. The comparison of composites images
before and after heating resulted that the heating caused the
formation of larger particles on the surface (Fig. 2c with d
and f with g). Inasmuch as it has not been added any silver
particle to the system during the heating process, the for-
mation of these particles is caused due to adhesion the
loaded silver on the surface of SG during the molten salt
process. In other words, during the process of molten salt at
temperatures and times of utilized in this research, particles
that formed on the surface had not enough time to move on
the SG and join together and form larger particles. The
heating of composites prepares this situation. Another point
is the size and the amount of formed nanoparticles. In
reference to Fig. 2d and e and two samples, which prepared
at 300 �C, the increasing of the time of molten salt process
caused to the formation of larger nanoparticles on the
surface. These results correspond to the changes of formal
colors of the composites (Fig. 1).
For the better investigation of the amount of fixed silver
on the SG in the composites, l-EDXRF was used (Fig. 3).
On the basis of the past researches, the intensity of the peak
of the plot for each of the elements is directly related to
their concentration [15]. The peck at the energy of 2.9 eV
represents silver in the composite. With due attention to
this energy and Fig. 3, in 300 and 150 �C, the amount of
silver in the composite has been increased with increasing
the time of molten salt process. On the other hand, at
constant time, the increasing of temperature causes the
same result. These results verify SEM results.
Water elution test
The results of water elution of silver/SG nanocomposites
are given in Fig. 4. This nanocomposite has been prepared
for bactericide action. The importance of this test is that
whether this nanocomposite causes water pollution or not.
On the contrary, the more stability of nanocomposites, the
more aged. This test exerts more intense condition to the
nanocomposites than industrial use condition. It is usually
utilized laminar flow in industrial systems; therefore, it
exerts less tension than turbulent flow on the nanocom-
posites which used in this experiment. As Fig. 4 indicates,
the amount of releasing silver from all of the nanocom-
posites is negligible. On the other hand, during the first 2 h,
the most amount of silver is released, but after this period
the releasing of silver is decreased so that the amount of
silver after 6 h becomes constant. Thus, one can claim that
the stability of prepared nanocomposite is suitable. Also
the increasing of time and temperature caused the
increasing of released silver. The reason is mounted silver
on the support, which has not strong binding with the
support. It may be eluted better from the composites after
preparation and this causes elution of these particles and
decreases the released silver.
Antibacterial test
Table 1 shows the antibacterial activity of the nanocom-
posites against E. coli and S. aureus. Generally, all silver-
doped products showed antibacterial activity, while the
parent SG has no inhibitory effect. It can be seen that
inhibition of sample prepared in 150 �C is less than
Fig. 1 The color of silver/silica gel nanocomposite at various temperatures and times
J Nanostruct Chem (2016) 6:215–221 217
123
prepared samples in 300 �C (Table 1). In particular, for
prepared samples in 300 �C, the zone of inhibition is
almost similar for all silver-doped products.
The results of this research are comparable with the
results of Hilonga et al. [4] study and our research has
higher antibacterial effects relatively. They prepared the
Ag/SG nanocomposite using the sol-gel method and in
several steps, however, their method is time consuming
and more complicated. In 2011, Quang et al. [5] pre-
pared Ag/SG nanocomposite via chemical reduction of
silver nitrate and the obtained hallow against E. coli was
equal to 6 mm. Referring to Table 1, the prepared
samples in our study at 300 �C had hollows about 6 mm
against E. coli.
Fig. 2 The SEM micrographs of parent silica gel (a, b) and silver/silica gel nanocomposites prepared at 150 �C (c) and 300 �C (d) for 30 min
then samples (c, d) annealed at 100 �C for 3 h (e, f), respectively
218 J Nanostruct Chem (2016) 6:215–221
123
Fig. 3 The l-EDXRF spectrum of composites, at deposition times of a 1 min and b 30 min at 100 �C and c 1 min and d 30 min at 300 �C
J Nanostruct Chem (2016) 6:215–221 219
123
Conclusions
An easy and new method was developed for the prepara-
tion of a bactericide nanocomposite. For the synthesis of
the nanocomposite, SG was introduced into the molten salt
of silver nitrate. After preparation of the nanocomposite,
SEM and l-EDXRF were utilized for its evaluation. The
nanocomposite showed antibacterial properties. The pre-
pared nanocomposite was stable during elution with water.
There was negligible released silver during elution with
water.
Open Access This article is distributed under the terms of the
Creative Commons Attribution 4.0 International License (http://crea
tivecommons.org/licenses/by/4.0/), which permits unrestricted use,
distribution, and reproduction in any medium, provided you give
appropriate credit to the original author(s) and the source, provide a
link to the Creative Commons license, and indicate if changes were
made.
References
1. Girase, B., Depan, D., Shah, J.S., Xu, W., Misra, R.D.K.: Silver–
clay nanohybrid structure for effective and diffusion-controlled
antimicrobial activity. Mater. Sci. Eng. C 31, 1759–1766 (2011)
2. Ghorbanpour, M.: Optimization of sensitivity and stability of
gold/silver bi-layer thin films used in surface plasmon resonance
chips. J. Nanostruct. 3, 309–313 (2013)
3. Rai, M., Yadav, A., Gade, A.: Silver nanoparticles as a new
generation of antimicrobials. Biotechnol. Adv. 27, 76–83 (2009)
4. Hilonga, A., Kim, J.K., Sarawade, P.B., Quang, D.V., Shao, G.,
Elineema, G., Taik Kim, H.: Silver-doped silica powder with
antibacterial properties. Powder Technol. 215–216, 219–222 (2012)
5. Quang, D.V., Sarawade, P.B., Hilonga, A., Kim, J.K., Chai, Y.G.,
Kim, S.H., Ryu, J.Y., Taik Kim, H.: Preparation of silver
nanoparticle containing silica micro beads and investigation of
their antibacterial activity. Appl. Surf. Sci. 257, 6963–6970
(2011)
6. Richardson, S., Postigo, C.: Drinking water disinfection by-
products. In: The Handbook of Environmental Chemistry (2012)
7. Cao, G.F., Sun, Y., Chen, J.G., Song, L.P., Jiang, J.Q., Liu, Z.T.,
Liu, Z.W.: Sutures modified by silver-loaded montmorillonite
with antibacterial properties. Appl. Clay Sci. 93–94, 102–106
(2014)
8. Rivera-Garza, M., Olguon, M.T., Garcoa-Sosa, I., Alcantara, D.,
Rodroguez Fuentes, G.: Silver supported on natural Mexican
zeolite as an antibacterial material. Microporous Mesoporous
Mater. 39, 431–444 (2000)
9. Ghorbanpour, M.: Stability modification of SPR silver nano-chips
by alkaline condensation of aminopropyltriethoxysilane.
J. Nanostruct. 5, 105–110 (2015)
10. Varma, R.S., Kothari, D.C., Tewari, R.: Nano-composite soda
lime silicate glass prepared using silver ion exchange. J. Non-
Cryst. Solids 355, 1246–1251 (2009)
11. Verne, E., Di Nunzio, S., Bosetti, M., Appendino, P., Vitale
Brovarone, C., Maina, G., Cannas, M.: Surface characterization
of silver-doped bioactive glass. Biomaterials 26, 5111–5119
(2005)
12. Xu, K., Wang, J.X., Kang, X.L., Chen, J.F.: Fabrication of
antibacterial monodispersed Ag-SiO2 core-shell nanoparticles
with high concentration. Mater. Lett. 63, 31–33 (2009)
0
0.2
0.4
0.6
0.8
1
1.2
1.4
1.6
0 5 10 15 20
Silv
er c
once
ntra
tion
(ppm
)
Time (h)
1 min, 150 °C
1 min, 300 °C
5 min, 150 °C
5 min, 300 °C
Fig. 4 The release of silver into
deionized water after a 24 h
period at 37 �C
Table 1 Antibacterial activity of composites
Temperature of
process (�C)
Time of
process (min)
Inhibition zones (mm)
E. coli S. aureus
150 1 0.7 ± 0.2 2.82 ± 0.6
5 0.4 ± 0.2 2.05 ± 0.3
10 0.9 ± 0.4 3.33 ± 0.4
20 2.5 ± 1.1 3.10 ± 0.3
30 4.1 ± 0.7 3.23 ± 0.8
300 1 6.0 ± 1.0 2.95 ± 1.1
5 6.3 ± 1.4 7.07 ± 1.3
10 5.85 ± 1.3 6.68 ± 1.5
20 5.9 ± 1.7 6.49 ± 1.2
30 5.7 ± 1.5 5.24 ± 2.0
220 J Nanostruct Chem (2016) 6:215–221
123
13. Zhang, X., Niu, H., Yan, J., Cai, Y.: Immobilizing silver
nanoparticles onto the surface of magnetic silica composite to
prepare magnetic disinfectant with enhanced stability and
antibacterial activity. Colloids Surf. A 375, 186–192 (2011)
14. Parandhaman, T., Das, A., Ramalingam, B., Samanta, D., Sastry,
T.P., Mandal, A.B., Das, S.K.: Antimicrobial behavior of
biosynthesized silica–silver nanocomposite for disinfection of
water: a mechanistic perspective. J. Hazard. Mater. 290, 117–126
(2015)
15. Ghorbanpour, M., Falamaki, C.: Micro energy dispersive X-ray
fluorescence as a powerful complementary technique for the
analysis of bimetallic Au/Ag/glass nanolayer composites used in
surface plasmon resonance sensors. Appl. Opt. 51, 7733–7738
(2012)
16. Ghorbanpour, M., Falamaki, C.: A novel method for the pro-
duction of highly adherent Au layers on glass substrates used in
surface plasmon resonance analysis: substitution of Cr or Ti
intermediate layers with Ag layer followed by an optimal
annealing treatment. J. Nanostruct. Chem. 3, 661–667 (2013)
J Nanostruct Chem (2016) 6:215–221 221
123